PROJECT SUMMARY Little is known about the role of micromechanical stress in regulating neuronal morphological and functional polarity. Diffuse axonal injury caused by mechanical impact displays characteristic axonal varicosities (swelling or beading), which are a prominent feature of traumatic brain injury (TBI). Abundant axonal varicosities are also a key sign for irreversible neurodegeneration in Alzheimer's and Parkinson's diseases, and multiple sclerosis. Under physiological conditions, a lower level of axonal varicosities can be observed in the brain. Although axonal varicosities profoundly affect action potential propagation and synaptic transmission, how they are specifically induced in axons by mechanical stress and regulated in health and disease remains a mystery. Our preliminary studies have led to several novel findings to shed light on this important question. We found that mechanical stress induces varicosity formation in unmyelinated axons, but not in dendrites or myelinated axons of central neurons. This process is unexpectedly rapid and reversible, where a transient receptor potential (TRP) channel acts as the major mechanosensitive (MS) ion channel. We further identified a novel binding protein of this channel, which regulates microtubule (MT) disassembly in response to Ca2+ influx. Moreover, we observed the rapid development of axonal varicosities in the brain of a mouse model of mild TBI. Based on our preliminary results, we propose an original hypothesis that micromechanical stress preferentially induces axonal varicosities in central neurons, and this process is regulated by axonal intrinsic and extrinsic mechanisms. To test this hypothesis, we will use a multidisciplinary approach including novel microbiomechanical assays, protein biochemistry, electrophysiological recording, state-of-the- art imaging, myelin coculture, knockout mice and a mild TBI mouse model. We will determine (Aim 1) whether targeting and activity of MS ion channels regulate polarized mechanosensation in central neurons, (Aim 2) how MT disassembly is induced by intra-axonal Ca2+ increase and in turn leads to varicosity formation, and (Aim 3) whether the pattern of axonal varicosity formation in the mild TBI mouse model is regulated by myelin, the MS channel and its binding protein. This project represents an underexplored research field with many open questions. This research is significant because it will provide novel mechanistic insights into a new form of central neuron polarity, polarized mechanosensation.